Objective Recombination events are common and serve as the primary driving force of diverse human adenovirus(HAdV),particularly in children with acute respiratory tract infections(ARIs).Therefore,continual monitoring ...Objective Recombination events are common and serve as the primary driving force of diverse human adenovirus(HAdV),particularly in children with acute respiratory tract infections(ARIs).Therefore,continual monitoring of these events is essential for effective viral surveillance and control.Methods Respiratory specimens were collected from children with ARIs between January 2022 and December 2023.The penton base,hexon,and fiber genes were amplified from HAdV-positive specimens and sequenced to determine the virus type.In cases with inconsistent typing results,genes were cloned into the pGEM-T vector to detect recombination events.Metagenomic next-generation sequencing(mNGS)was performed to characterize the recombinant HAdV genomes.Results Among 6,771 specimens,277(4.09%,277/6,771)were positvie for HAdV,of which 157(56.68%,157/277)were successfully typed,with HAdV-B3 being the dominant type(91.08%,143/157),and 14(5.05%,14/277)exhibited inconsistent typing results,six of which belonged to species B.The penton base genes of these six specimens were classified as HAdV-B7,whereas their hexon and fiber genes were classified as HAdV-B3,resulting in a recombinant genotype designated P7H3F3,which closely resembled HAdV-B114.Additionally,a partial gene encoding L152/55 kD was identified,which originated from HAdV-B16.Conclusion A novel recombinant,P7H3F3,was identified,containing sequences derived from HAdV-B3 and HAdV-B7,which is similar to HAdV-B114,along with additional sequences from HAdV-B16.展开更多
OBJECTIVE： To test the in vitro antiviral activity of a crude tissue extract （CTE/from the earthworm Eisenia fetida, determine any effective components in the CTE, andelucidate possiblemechanismsofaction. METHODS： ...OBJECTIVE： To test the in vitro antiviral activity of a crude tissue extract （CTE/from the earthworm Eisenia fetida, determine any effective components in the CTE, andelucidate possiblemechanismsofaction. METHODS： A CTE was made by homogenizing earthworms, followed by treatment with ammoni- um sulfate, then thermal denaturation. Inhibition of virus-induced cytopathic effect （CPE） was used to assess antiviral activity. Chromatographic analy- sis was used to identify effective components in the CTE. RESULTS： The CTE inhibited viral CPE at non-cyto- toxic concentrations. Chromatography indicated that antiviral components corresponded to three active peaks indicative of proteases, nucleases and lysozymes. For adenoviruses, reduction in viral ac- tivity occurred for 100 lag/mL CTE. The reduction in adenoviral activity for four fractions was 100%, 91.8%, 86.9%, and 94.7%. For influenza viruses, re- duction in viral activity of 100%, 86.6%, 69.1% and 88.3% was observed for 37 pg/mL CTE. In addition, three active fractions mixture had stronger antiviral activity （98.7% and 96.7%） than three fractions alone.Gel electrophoresis results indicated that nu- cleases from E. fetida could degrade the genome of influenza viruses and adenoviruses. CONCLUSION： The earthworm CTE displayed non-specific antiviral properties, possibly mediated by a combination of proteases, nucleases and lyso- zymes. Nucleases likely participate in the antiviral process, and degrade the genome of the virus thereby preventing further replication.展开更多
To find a technique of detecting and differentiating enteric adenoviruses (EAds) in clinical samples, a novel PCR approach was developed. EAds were able to be detected by use of a pair of subgroup F general primers (...To find a technique of detecting and differentiating enteric adenoviruses (EAds) in clinical samples, a novel PCR approach was developed. EAds were able to be detected by use of a pair of subgroup F general primers (P1 and P2), and they were also be able to be differentiated from each other in the presence of another adenovirus type 40 (Ad40) specific primer (P3) in the same tube. Our results showed that there was one band for Ad41 and two bands for Ad40, respectively, on running-gel after PCR performance. PCR was performed on 40 specimens in parallel directly with dot-hybridization assay on the same diluted stool samples. 20 of 40 specimens were positive by hybridization (of them 12 were Ad41 and 8 were Ad40), whereas 26 were positive by PCR performance on the same samples with Ad41 18 and Ad40 8 positive as well. Our study indicated that this novel method could be used in clinical laboratory or in epidemic investigation for Eads展开更多
Interleukin-2(IL-2) has been demonstrated to beone of the most effective target genes in cancerimmunogene therapy. There are more than 20 clinicalprotocols of cancer gene therapy introducing IL-2 intotumor patients to...Interleukin-2(IL-2) has been demonstrated to beone of the most effective target genes in cancerimmunogene therapy. There are more than 20 clinicalprotocols of cancer gene therapy introducing IL-2 intotumor patients to treat melanoma, renal carcinoma,prostate carcinoma, colon carcinoma,展开更多
Recombinant adenoviruses encoding murine IL 2 gene or IL 3 gene were directly injected into established subcutaneous tumor model of G422 glioblastoma cells. After treatment, the tumor size and survival of the gliobl...Recombinant adenoviruses encoding murine IL 2 gene or IL 3 gene were directly injected into established subcutaneous tumor model of G422 glioblastoma cells. After treatment, the tumor size and survival of the glioblastoma bearing mice were observed. The splenic NK and CTL cytotoxicities were detected by standard 4 hour 51 Cr release assay. We also examined the histopathological changes of tumor by hematoxylin and eosin staining. The results showed that intratumoral injection of adenoviruses encoding murine IL 2 gene or IL 3 gene significantly inhibited the growth of G422 glioblastoma and prolonged the survival period of glioblastoma bearing mice. The CTL cytotoxicity of the gene therapy groups was significantly higher than that of the control groups, but NK activity remained unchanged, indicating that specific immunity contributes to the in vivo antitumor effect of the direct gene therapy. There were much more tumor necrosis and inflammatory cell infiltration in the tumor of the gene therapy groups. Combined IL 2/IL 3 gene therapy could induce higher level of CTL and enhance the therapeutic potential further. The results suggest that intratumoral injection of recombinant adenoviruses encoding certain kind of cytokines may be a useful approach in the treatment of a malignancy of the central nervous system.展开更多
Human adenoviruses(HAdVs)can cause acute hepatitis in immunocompromised patients.However,it is unclear whether HAdVs are contributors to hepatitis in immunocompetent children.In this study,the liver function test(LFT)...Human adenoviruses(HAdVs)can cause acute hepatitis in immunocompromised patients.However,it is unclear whether HAdVs are contributors to hepatitis in immunocompetent children.In this study,the liver function test(LFT)results were retrospectively analyzed among children hospitalized(age<14 years)between January 2016 and October 2019 for acute respiratory infection caused by adenoviruses.Alanine transaminase(ALT)and aspartate aminotransferase(AST)levels were elevated in 7.74%and 46.89%of patients,respectively.All patients with>2 folds of the upper limit of ALT or AST levels were infected with HAdV-7 or HAdV-55.Significantly higher levels of ALT,AST,γ-glutamyl transpeptidase(γ-GT),and lower albumin levels were observed in the HAdV-7 infection group than in the HAdV-3 infection group.HAdV-55 infection led to significantly higherγ-GT,total bilirubin,and direct bilirubin levels than the other infection types.The records of four patients with serial monitoring of the LFT results were further analyzed.Multiple indicators remained abnormal during the entire hospitalization in these patients.These results indicate that HAdV infection is often accompanied by abnormal liver function,and HAdV-7 and HAdV-55 might be under-recognized contributors to hepatitis among children.展开更多
Background Suicide gene therapy is a widely used molecular treatment for head and neck cancer. In this study, we try to use the method of homogenous recombination in bacteria to clone thymidine kinase gene （tk）-a ki...Background Suicide gene therapy is a widely used molecular treatment for head and neck cancer. In this study, we try to use the method of homogenous recombination in bacteria to clone thymidine kinase gene （tk）-a kind of suicide gene to adenovirus backbone vectors for the construction of replication-defective adenoviruses. Methods pAdTrack-CMV/tk was constructed through subclone of a restriction endonuclease fragment including thymidine kinase gene from plasmid pCMV-tk to another plasmid pAdTrack-CMV, and then co-transfected with supercoiled pAdEasy-1, which was an adenoviral backbone vector except for deletions of E1 and E3, to competent E. coli BJ5183 for homogenous recombination using electroporation procedure. With the same method, pAdTrack-CMV was also co-transformed with pAdEasy-1 for homogenous recombination in BJ5183. Identified with restriction endonuclease Pad and polymerase chain reaction （PCR）, plasmids pAd-GFP/tk and pAd-GFP were successfully constructed. Each of them was digested with Pacl and sequently transfected into human embryo kidney 293 cells （HEK293） using Lipofectamine 2000. Results Comet-like adenovirus-producing foci of Ad-GFP/tk and Ad-GFP were observed after 5 to 7 days of cell culture After twelve days of packaging, the replication-defective adenoviruses were collected. Identified with PCR, thymidine kinase gene was successfully constructed into Ad-GFP/tk. Conclusion The replication-defective adenoviruses containing thymidine kinase can be constructed more easily by homogenous recombination in bacteria than conventional techniques.展开更多
Glioblastoma Multiforme(GBM)is a rapidly progressing brain tumor.Despite the relatively low percentage of cancer patients with glioma diagnoses,recent statistics indicate that the number of glioma patients may have in...Glioblastoma Multiforme(GBM)is a rapidly progressing brain tumor.Despite the relatively low percentage of cancer patients with glioma diagnoses,recent statistics indicate that the number of glioma patients may have increased over the past decade.Current therapeutic options for glioma patients include tumor resection,chemotherapy,and concomitant radiation therapy with an average survival of approximately 16 months.The rapid progression of gliomas has spurred the development of novel treatment options,such as cancer gene therapy and oncolytic virotherapy.Preclinical testing of oncolytic adenoviruses using glioma models revealed both positive and negative sides of the virotherapy approach.Here we present a detailed overview of the glioma virotherapy field and discuss auxiliary therapeutic strategies with the potential for augmenting clinical efficacy of GBM virotherapy treatment.展开更多
Objective:To investigate the anti-tumor effects of an E1B55KD-deleted oncolytic adenovirus,H101,in combination with a humanized anti-PD-1(Programmed cell death protein 1)monoclonal antibody,Camrelizumab.Methods:Anti-t...Objective:To investigate the anti-tumor effects of an E1B55KD-deleted oncolytic adenovirus,H101,in combination with a humanized anti-PD-1(Programmed cell death protein 1)monoclonal antibody,Camrelizumab.Methods:Anti-tumor efficacy of intratumoral injection of H101 or/and intraperitoneal injection of Camrelizumab were evaluated in an immune system humanized NOD Prkdc^(scid) Il2rg^(-/-)mice subcutaneous(S.C.)tumor model,established with human glioblastoma of unknown origin cell line U87-MG,and human bladder cancer cell line T24 and YTS-1.The mechanism by which H101 induced anti-tumor immunity were also investigated.Results:Combining H101 with Camrelizumab demonstrated more potent anti-tumor effects than monotherapy in mouse S.C.tumor model.Increased tumor-infiltrating T cells were observed in the combined treatment group.H101 infection decreased the expression of CD47 in cancer cells,thereby promoting macrophages to phagocytose cancer cells.Following the H101-mediated activation of macrophages,increased levels of cytokines,including TNF,IL-12 and IFN-γwere observed.Moreover,when induced THP-1 cells were co-cultured with H101-treated cancer cells,expression of IFN-γwas increased in T cells.Elimination of IL-12 using an anti-IL-12 antibody abolished IFN-γproduction from T cells.In addition,infection with H101 increased PD-L1 expression in YTS-1 cells.These results suggested that H101 may act synergistically to enhance the therapeutic efficacy of PD-1 blockade in cancer via suppressing CD47 signaling,which may promote macrophages to phagocytose tumor cells and activate CD8^(+)T cells.Conclusion:The combination of H101 with PD-1 blockade exhibits potential as a novel strategy for the treatment of cancer.展开更多
BACKGROUND Type 1 diabetes(T1D)results from the autoimmune-mediated loss of pancreatic β-cells.Current insulin therapies offer symptomatic relief but fall short of providing a definitive cure.Islet cell transplantati...BACKGROUND Type 1 diabetes(T1D)results from the autoimmune-mediated loss of pancreatic β-cells.Current insulin therapies offer symptomatic relief but fall short of providing a definitive cure.Islet cell transplantation,while promising,faces limitations related to donor scarcity,procedural complexities,and the necessity for long-term immunosuppression.Consequently,there is an urgent need for innovative strategies aimed at β-cell regeneration.Patient-derived induced pluripotent stem cells(iPSCs),obtained from peripheral blood mononuclear cells(PBMCs)of T1D patients,hold great potential as a source of cells for therapeutic purposes.Therefore,the differentiation of T1D-iPSCs into functional pancreatic β-cells is a critical step toward effective β-cell replacement therapy.AIM To assess the potential of patient-derived T1D-β-like cells(differentiated from T1D-iPSCs reprogrammed from T1D-PBMCs)for restoring β-cell function in T1D.METHODS T1D-iPSCs were reprogrammed from T1D-PBMCs using an episomal vectorbased approach.Pluripotency was confirmed by flow cytometry(FCM),quantitative real-time polymerase chain reaction,genomic stability analysis,and teratoma formation assays.Differentiation into pancreatic β-cells was optimized using triiodothyronine(T3),vitamin C(Vc),and an adenovirus(M3C)encoding pancreatic duodenal homeobox-1,neurogenin 3(Ngn3),and MAF bZIP transcription factor A(MafA).Following characterization of β-cell features by immunofluorescence,quantitative real-time polymerase chain reaction,and flow cytometry,therapeutic efficacy was assessed through blood glucose monitoring after transplantation under the renal capsule of streptozotocin-induced diabetic mice.RESULTS T1D-iPSCs were successfully generated from T1D-PBMCs.These cells exhibited the hallmark characteristics of pluripotent stem cells,including appropriate morphology,differentiation potential,genomic integrity,and expression of pluripotency-associated genes.Differentiation yielded insulin-positive(insulin+)pancreatic β-like cells that,at the mRNA level,expressed key β-cell markers such as pancreatic duodenal homeobox-1,Ngn3,MafA,NeuroD,glucagon-like peptide-1 receptor,Nkx6.1,glucose transporter 2,and Kir6.2.Notably,the T3+Vc group displayed the lowest Ngn3 expression(1.31±0.38 vs 1.96±0.25 vs 2.51±0.24,P<0.01),while the M3C+T3+Vc group exhibited the highest MafA expression(0.49±0.11 vs 0.32±0.06 vs 0.29±0.08,P<0.05).Both in vitro and in vivo assessments confirmed the insulin secretion ability of the generated β-like cells;however,they did not demonstrate appropriate modulation of insulin release in response to variations in extracellular glucose concentrations.CONCLUSION T1D-iPSCs derived from T1D-PBMCs can be differentiated into insulin+β-like cells,albeit with functional immaturity.These cells represent a potential source of seed cells for β-cell replacement therapy in T1D.展开更多
Fowl adenovirus(FAdV)serotype 4,recognized as the causative agent of hydropericardium syndrome(HPS)in chickens,causes substantial economic losses in poultry farming.To develop a simple,rapid,and reliable diagnostic me...Fowl adenovirus(FAdV)serotype 4,recognized as the causative agent of hydropericardium syndrome(HPS)in chickens,causes substantial economic losses in poultry farming.To develop a simple,rapid,and reliable diagnostic method for the timely detection of FAdV-4 nucleic acid,we integrated the CRISPR/Cas12a system with recombinase-aided amplification(RAA).This approach enables visual detection of FAdV-4 with a sensitivity of one genome copy.The results can be obtained within 40 to 50 min without the need for complex instrumentation,making it ideal for remote field applications.Using this method,we investigated the prevalence of FAdV-4 in both common farm poultry and wild birds.Our results indicated that the FAdV-4-positive rate in wild birds was 51.19%,suggesting that wild birds may serve as specific reservoirs for this virus.In summary,we present a sensitive,swift,accurate,and inexpensive detection method for FAdV-4,along with an investigation of its epidemic situation in birds.Our study advances the detection and epidemiological understanding of FAdV-4 transmission among farm poultry and wild birds.展开更多
Acute respiratory tract infections(ARTIs)are among the leading causes of morbidity and mortality in children worldwide.Human adenovirus(HAdV)infections are estimated to account for at least 5%of pediatric ARTIs.The ci...Acute respiratory tract infections(ARTIs)are among the leading causes of morbidity and mortality in children worldwide.Human adenovirus(HAdV)infections are estimated to account for at least 5%of pediatric ARTIs.The circulated genotypes of HAdV and the correlation between genotype and clinical manifestations in Wuhan,China,before and after the complete relaxation of nonpharmaceutical interventions against severe acute respiratory syndrome coronavirus 2,remain unknown.Here,101 HAdV strains were isolated from throat swab samples collected from hospitalized children with ARTIs who tested positive for HAdV nucleic acid.Of these,sixty-six strains from 2022 to twenty-three strains from 2023 were successfully genotyped and subjected to phylogenetic analysis based on the hexon,penton base,and fiber genes.Six genotypes,B3,C1,C2,C5,C104,and C108 were identified.HAdV-B3(84.85%)was the most prevalent type in 2022,while HAdV-C(86.96%),including C1,C2,C108,and C104,was the most prevalent in 2023.These strains were phylogenetically related to strains from Japan,China,and the United States in recent years.When comparing clinical characteristics,pediatric patients infected with B3,C1,C2,C5,C104,or C108 exhibited similar clinical manifestations,primarily fever and cough,but varying interleukin(IL)-10 levels.In conclusion,from June 2022 to September 2023,the circulated genotypes of HAdV in Wuhan included B3,C1,C2,C108,C5,and C104.The endemic pattern of HAdV in Wuhan,China,shifted from species B as the dominant type in 2022 to species C in 2023.展开更多
OBJECTIVE:To investigate the mechanism of Dan Ze mixture(丹泽合剂,DZM)in the treatment of lipotoxic cardiomyopathy.METHODS:Ultra-performance liquid chromatography tandem mass spectrometry was employed to characterize ...OBJECTIVE:To investigate the mechanism of Dan Ze mixture(丹泽合剂,DZM)in the treatment of lipotoxic cardiomyopathy.METHODS:Ultra-performance liquid chromatography tandem mass spectrometry was employed to characterize the serum migration constituents of DZM.A lipotoxic cardiomyopathy rat model was established through high-fat diet and intervened by different doses of DZM.The cardiac function was assessed using echocardiography,and hematoxylin and eosin,oil red O,and Masson staining were conducted to evaluate morphological changes,lipid accumulation,and fibrosis in myocardial tissue.Serum myocardial enzyme activity,lipid levels,and lipid content of myocardial tissue were measured,while fluorescent staining and colorimetry were used to assess oxidation levels in myocardial tissue.Mitochondrial membrane potential was detected by 5,5',6,6'-Tetrachloro-1,1',3,3'-tetraethyl-imidacarbocyanineiodide(JC-1).Transmission electron microscopy was employed to observe ultrastructure and mitochondrial structure changes in myocardial tissue.Fluorescence double staining and colocalization were utilized to observe the binding of autophagosomes and mitochondria,while immunohistochemical staining was used to detect the expression of mitophagy-related proteins.Terminal deoxynucleoitidyl transferase mediated nick end labeling staining was employed for the identification of apoptosis in myocardial tissue,while quantitative real-time reverse transcriptase polymerase chain reaction(q RT-PCR)and Western blot were utilized for the detection of apoptosis,B-cell lymphoma-2 adenovirus E1B 19 k Da-interacting protein 3(BNIP3)/mitophagy signaling pathway-related genes and proteins.In palmitic acid-induced Rat H9C2 cardiomyocytes(H9c2)cells,various cellular parameters including cell viability,lactate dehydrogenase release,apoptosis rate,oxidative stress level,mitochondrial structure and function,and mitophagy level were assessed after the treatment of DZM drug-containing serum for a duration of 24 h.The cellular expressions of BNIP3/mitophagy signaling pathway relevant genes and proteins were further evaluated using q RT-PCR and Western blot techniques.RESULTS:A total of 295 prototypes(e.g.,phenolic acids,quinones,terpenoids)were identified in serum of rats after oral administration of DZM.In vivo,DZM therapy has been shown to effectively enhance cardiac function,mitigate high-fat diet-induced myocardial structural damage and lipid accumulation.Furthermore,DZM has demonstrated the ability to reduce lipid levels,attenuate cell apoptosis,combat oxidative stress,enhance mitochondrial structure and function,and activate the BNIP3/mitophagy signaling pathway.Furthermore,the silencing of BNIP3 has been shown to exacerbate palmitic acid-induced damages in H9c2 cells,while inhibiting the BNIP3/mitophagy signaling pathway can mitigate the inhibitory effects of DZM on palmitic acidinduced apoptosis,lipid deposition and oxidative stress.CONCLUSION:This study presents preliminary evidence for the therapeutic efficacy of DZM on lipotoxic cardiomyopathy through the activating BNIP3/mitophagy signaling pathway.展开更多
Human adenovirus type 108(HAdV-108)has been detected in multiple countries,including China,and is associated with severe acute respiratory infection(ARI)in children,with reported fatalities.However,studies on HAdV-108...Human adenovirus type 108(HAdV-108)has been detected in multiple countries,including China,and is associated with severe acute respiratory infection(ARI)in children,with reported fatalities.However,studies on HAdV-108 remain limited.This study aimed to investigate the clinical and genetic characteristics of HAdV-108 in ARI children in China.From 2014 to 2024,6720 respiratory samples were collected from hospitalized children with ARI at ten hospitals across northern and southern China,of which 505(7.51%)tested positive for HAdV.The whole-genome and three major capsid protein genes were amplified and sequenced for bioinformatics analysis,which revealed that among 317 HAdV-isolated samples,21(6.62%)were identified as HAdV-108,ranking third after HAdV-114 and HAdV-7.Clinical analysis of HAdV-108-positive cases showed that the main manifestations were cough and fever.Seven children had gastrointestinal symptoms,and two children without underlying diseases were diagnosed with severe pneumonia.Phylogenetic analysis of wholegenome sequences revealed distinct predominant epidemic branches between domestic and international strains,with one strain obtained in this study forming an independent branch.Hexon protein exhibited the fastest evolution rate,lowest identity,and greatest amino acid variability,while fiber protein displayed the slowest evolution rate,highest identity,and greatest conservation and stability.Compared with the earliest reported HAdV-108 strain,three amino acid deletions were identified in the RGD loop region of penton base protein,resulting in potential structural change.Recombination analysis identified five distinct recombination patterns.In vitro experiments demonstrated that HAdV-108 had proliferation capacity comparable to other species C adenoviruses.In summary,HAdV-108 has persistently circulated in China,causing severe ARIs and concurrent gastrointestinal manifestations.Cluster3 was the predominant epidemic branch in China.HAdV-108 exhibited significant intratype genetic variation,with random and diverse recombination events.展开更多
BACKGROUND Congenital short bowel syndrome(CSBS)is a rare disorder characterized by a congenital shortage in the length of the small intestines,resulting in compromised intestinal functionality,frequently accompanied ...BACKGROUND Congenital short bowel syndrome(CSBS)is a rare disorder characterized by a congenital shortage in the length of the small intestines,resulting in compromised intestinal functionality,frequently accompanied by congenital intestinal malrotation.This study summarizes the experience of enteral and parenteral nutrition(PN)schemes for two cases of CSBS accompanied by intestinal malrotation to provide a reference for this condition.CASE SUMMARY Case 1 underwent surgical intervention 23 days after birth,but experienced postoperative intolerance to oral feeding.Consequently,the patient was transitioned to nasal feeding for 6 months while concurrently receiving round-the-clock PN.Despite ongoing adjustments to the caloric intake of enteral and PN throughout the treatment,cholestatic liver injury persisted.Furthermore,this child developed malnutrition by the age of 1 year.Case 2 received surgical intervention on 2 days after birth,demonstrating favorable tolerance for enteral feeding and intermittent PN.The child exhibited satisfactory growth and development without any associated complications.CONCLUSION CSBS is rarely encountered in clinical practice and is often accompanied by congenital intestinal malrotation.It can be improved with early diagnosis and active surgical intervention.Thus,we recommend early initiation of total PN,which is a highly effective measure in promoting early growth and development.展开更多
[Objective] Sheep contagious ecthyma virus B2L gene recombinant adenovirus was built by adenovirus vector system.[Method] Genome DNA extracted from sheep contagious ecthyma virus strain JLSY04 as a template,Gene fragm...[Objective] Sheep contagious ecthyma virus B2L gene recombinant adenovirus was built by adenovirus vector system.[Method] Genome DNA extracted from sheep contagious ecthyma virus strain JLSY04 as a template,Gene fragments obtained from B2L by PCR amplification;B2L gene cloning was cloned into PDNR-CMD vector,screening positive clones and plasmid CTC572-6 was obtained;CTC572-6 plasmid for homologous was recombined with the adenoviral vector.Screening positive clones and bacilli PCR,digestion and sequencing and so on were identified.[Result] After identified by enzyme digestion and gene sequencing,recombinant adenovirus vector CTC572Ade-30 of carrying sheep contagious ecthyma virus B2L gene was constructed successfully.[Conclusion] Which laid the foundation for sheep contagious ecthyma genetically engineered vaccine.展开更多
Objective: To evaluate the therapeutic efficacy of replicative adenovirus CNHK500 in the treatment of hepatocellular carcinoma. Methods: Virus proliferation assay, cell viability assay and Western blot were performed ...Objective: To evaluate the therapeutic efficacy of replicative adenovirus CNHK500 in the treatment of hepatocellular carcinoma. Methods: Virus proliferation assay, cell viability assay and Western blot were performed to assess the selective replication and cytolysis of CNHK500 in telomerase positive liver cancer cells Hep3B, HepGII, SMMC7721 and in normal cells. Results: The replicative multiples of CNHK500 in HepGII, Hep3B and SMMC7221 after 96 h of virus proliferation were 52 000, 396 984.9 and 632 911.3 fold respectively, similar to those of wtAd5. However, CNHK500 demonstrated more significant attenuated replicative ability in normal cell lines than wtAd5. CNHK500 replicated only 3.1-100 fold at 96 h, while the wtAd5 still reached 3160-17 357 fold. CNHK500 could cause half of HepGII cells death within 7 days at MOI 2, in Hep3B cell lines the IC50 was as low as MOI 0.01, whereas the IC50 in BJ cell was as high as MOI 1000. CNHK500 E1A protein could only be detected in hepatocellular cancer cells but not in normal cells under normoxia. E1B protein could only be detected under hypoxia condition at a MOI of 1. Conclusion: CNHK500 can efficiently replicate in and kill liver cancer cells as well as wtAd5 do while it is severely attenuated in proliferation and cytolysis among normal cells. It would be a prominsing strategy for liver cancer tratment.展开更多
Human adenoviruses (HAdVs) belong to the genus Mastadenovirus, family Adenoviridae, and can cause respiratory tract and gastrointestinal tract infections, as well as ocular infections in humans[1]. Till date, a total ...Human adenoviruses (HAdVs) belong to the genus Mastadenovirus, family Adenoviridae, and can cause respiratory tract and gastrointestinal tract infections, as well as ocular infections in humans[1]. Till date, a total of 84 unique genotypes of AdVs have been identified and classified as human Mastadenovirus species A to G, and specific types are often associated with certain clinical symptoms, epidemiological settings, and demographic risk groups[2]. Among these species, members of the species HAdV-B (HAdV-3, HAdV-7, HAdV-11, HAdV-16, HAdV-21, HAdV-34, HAdV-35, HAdV-50, etc), HAdV-C (HAdV-1, HAdV-2, HAdV-5, and HAdV-6), and HAdV-E (HAdV-4) have been generally associated with respiratory infections[3]. HAdV infections are mild and self-limited in healthy individuals, whereas they can result in high mortality rates in immunocompetent and immunocompromized patients[4].展开更多
AIM:To generate an adenoviral vector specifically targeting the EphA2 receptor(EphA2R) highly expressed on pancreatic cancer cells in vivo.METHODS:YSA,a small peptide ligand that binds the EphA2R with high affinity,wa...AIM:To generate an adenoviral vector specifically targeting the EphA2 receptor(EphA2R) highly expressed on pancreatic cancer cells in vivo.METHODS:YSA,a small peptide ligand that binds the EphA2R with high affinity,was inserted into the HI loop of the adenovirus serotype 5 fiber knob.To further increase the specificity of this vector,binding sites for native adenoviral receptors,the coxsackie and adenovirus receptor(CAR) and integrin,were ablated from the viral capsid.The ablated retargeted adenoviral vector was produced on 293T cells.Specifi c targeting of this novel adenoviral vector to pancreatic cancer was investigated on established human pancreatic cancer cell lines.Upon demonstrating specifi c in vitro targeting,in vivo targeting to subcutaneous growing human pancreatic cancer was tested by intravenous and intraperitoneal administration of the ablated adenoviral vector.RESULTS:Ablation of native cellular binding sites reduced adenoviral transduction at least 100-fold.Insertion of the YSA peptide in the HI loop restored adenoviral transduction of EphA2R-expressing cells but not of cells lacking this receptor.YSA-mediated transduction was inhibited by addition of synthetic YSA peptide.The transduction specificity of the ablated retargeted vector towards human pancreatic cancer cells was enhanced almost 10-fold in vitro.In a subsequent in vivo study in a nude(nu/nu) mouse model however,no increased adenoviral targeting to subcutaneously growing human pancreas cancer nodules was seen upon injection into the tail vein,nor upon injection into the peritoneum.CONCLUSION:Targeting the EphA2 receptor increases specificity of adenoviral transduction of human pancreatic cancer cells in vitro but fails to enhance pancreatic cancer transduction in vivo.展开更多
基金supported by the Capital’s Funds for Health Improvement and Research(CFH,shoufa-2022-1G-1131 and shoufa 2022-4G-1133)the High Level Technical Talent Construction Project of Beijing Municipal Health Commission(Discipline Leader-02-20)+1 种基金Beijing Municipal Public Welfare Development and Reform Pilot Project for Medical Research Institutes(JYY2023-10)the Pathogen Spectrum and Host Marker Analysis in Children with Respiratory Tract Infections of Children(Grant 2024-0040).
文摘Objective Recombination events are common and serve as the primary driving force of diverse human adenovirus(HAdV),particularly in children with acute respiratory tract infections(ARIs).Therefore,continual monitoring of these events is essential for effective viral surveillance and control.Methods Respiratory specimens were collected from children with ARIs between January 2022 and December 2023.The penton base,hexon,and fiber genes were amplified from HAdV-positive specimens and sequenced to determine the virus type.In cases with inconsistent typing results,genes were cloned into the pGEM-T vector to detect recombination events.Metagenomic next-generation sequencing(mNGS)was performed to characterize the recombinant HAdV genomes.Results Among 6,771 specimens,277(4.09%,277/6,771)were positvie for HAdV,of which 157(56.68%,157/277)were successfully typed,with HAdV-B3 being the dominant type(91.08%,143/157),and 14(5.05%,14/277)exhibited inconsistent typing results,six of which belonged to species B.The penton base genes of these six specimens were classified as HAdV-B7,whereas their hexon and fiber genes were classified as HAdV-B3,resulting in a recombinant genotype designated P7H3F3,which closely resembled HAdV-B114.Additionally,a partial gene encoding L152/55 kD was identified,which originated from HAdV-B16.Conclusion A novel recombinant,P7H3F3,was identified,containing sequences derived from HAdV-B3 and HAdV-B7,which is similar to HAdV-B114,along with additional sequences from HAdV-B16.
文摘OBJECTIVE： To test the in vitro antiviral activity of a crude tissue extract （CTE/from the earthworm Eisenia fetida, determine any effective components in the CTE, andelucidate possiblemechanismsofaction. METHODS： A CTE was made by homogenizing earthworms, followed by treatment with ammoni- um sulfate, then thermal denaturation. Inhibition of virus-induced cytopathic effect （CPE） was used to assess antiviral activity. Chromatographic analy- sis was used to identify effective components in the CTE. RESULTS： The CTE inhibited viral CPE at non-cyto- toxic concentrations. Chromatography indicated that antiviral components corresponded to three active peaks indicative of proteases, nucleases and lysozymes. For adenoviruses, reduction in viral ac- tivity occurred for 100 lag/mL CTE. The reduction in adenoviral activity for four fractions was 100%, 91.8%, 86.9%, and 94.7%. For influenza viruses, re- duction in viral activity of 100%, 86.6%, 69.1% and 88.3% was observed for 37 pg/mL CTE. In addition, three active fractions mixture had stronger antiviral activity （98.7% and 96.7%） than three fractions alone.Gel electrophoresis results indicated that nu- cleases from E. fetida could degrade the genome of influenza viruses and adenoviruses. CONCLUSION： The earthworm CTE displayed non-specific antiviral properties, possibly mediated by a combination of proteases, nucleases and lyso- zymes. Nucleases likely participate in the antiviral process, and degrade the genome of the virus thereby preventing further replication.
基金This project was supported by a grant from National Sci-ences Foundation of China( Serial No.3 93 70 0 3 8) and part ofa grant from Hubei Science and Technology Comm ittee( No.89JJ13 - 18)
文摘To find a technique of detecting and differentiating enteric adenoviruses (EAds) in clinical samples, a novel PCR approach was developed. EAds were able to be detected by use of a pair of subgroup F general primers (P1 and P2), and they were also be able to be differentiated from each other in the presence of another adenovirus type 40 (Ad40) specific primer (P3) in the same tube. Our results showed that there was one band for Ad41 and two bands for Ad40, respectively, on running-gel after PCR performance. PCR was performed on 40 specimens in parallel directly with dot-hybridization assay on the same diluted stool samples. 20 of 40 specimens were positive by hybridization (of them 12 were Ad41 and 8 were Ad40), whereas 26 were positive by PCR performance on the same samples with Ad41 18 and Ad40 8 positive as well. Our study indicated that this novel method could be used in clinical laboratory or in epidemic investigation for Eads
文摘Interleukin-2(IL-2) has been demonstrated to beone of the most effective target genes in cancerimmunogene therapy. There are more than 20 clinicalprotocols of cancer gene therapy introducing IL-2 intotumor patients to treat melanoma, renal carcinoma,prostate carcinoma, colon carcinoma,
文摘Recombinant adenoviruses encoding murine IL 2 gene or IL 3 gene were directly injected into established subcutaneous tumor model of G422 glioblastoma cells. After treatment, the tumor size and survival of the glioblastoma bearing mice were observed. The splenic NK and CTL cytotoxicities were detected by standard 4 hour 51 Cr release assay. We also examined the histopathological changes of tumor by hematoxylin and eosin staining. The results showed that intratumoral injection of adenoviruses encoding murine IL 2 gene or IL 3 gene significantly inhibited the growth of G422 glioblastoma and prolonged the survival period of glioblastoma bearing mice. The CTL cytotoxicity of the gene therapy groups was significantly higher than that of the control groups, but NK activity remained unchanged, indicating that specific immunity contributes to the in vivo antitumor effect of the direct gene therapy. There were much more tumor necrosis and inflammatory cell infiltration in the tumor of the gene therapy groups. Combined IL 2/IL 3 gene therapy could induce higher level of CTL and enhance the therapeutic potential further. The results suggest that intratumoral injection of recombinant adenoviruses encoding certain kind of cytokines may be a useful approach in the treatment of a malignancy of the central nervous system.
基金This study was supported by the National Natural Science Foundation of China(82072264,81970003)Natural Science Foundation of Guangdong Province,China(2021A1515011071)The study sponsors had no involvement in the study design,in the collection,analysis and interpretation of data,in the writing of the manuscript,and in the decision to submit the manuscript for publication.
文摘Human adenoviruses(HAdVs)can cause acute hepatitis in immunocompromised patients.However,it is unclear whether HAdVs are contributors to hepatitis in immunocompetent children.In this study,the liver function test(LFT)results were retrospectively analyzed among children hospitalized(age<14 years)between January 2016 and October 2019 for acute respiratory infection caused by adenoviruses.Alanine transaminase(ALT)and aspartate aminotransferase(AST)levels were elevated in 7.74%and 46.89%of patients,respectively.All patients with>2 folds of the upper limit of ALT or AST levels were infected with HAdV-7 or HAdV-55.Significantly higher levels of ALT,AST,γ-glutamyl transpeptidase(γ-GT),and lower albumin levels were observed in the HAdV-7 infection group than in the HAdV-3 infection group.HAdV-55 infection led to significantly higherγ-GT,total bilirubin,and direct bilirubin levels than the other infection types.The records of four patients with serial monitoring of the LFT results were further analyzed.Multiple indicators remained abnormal during the entire hospitalization in these patients.These results indicate that HAdV infection is often accompanied by abnormal liver function,and HAdV-7 and HAdV-55 might be under-recognized contributors to hepatitis among children.
基金the National Natural Science Foundation of China(No.30070808).
文摘Background Suicide gene therapy is a widely used molecular treatment for head and neck cancer. In this study, we try to use the method of homogenous recombination in bacteria to clone thymidine kinase gene （tk）-a kind of suicide gene to adenovirus backbone vectors for the construction of replication-defective adenoviruses. Methods pAdTrack-CMV/tk was constructed through subclone of a restriction endonuclease fragment including thymidine kinase gene from plasmid pCMV-tk to another plasmid pAdTrack-CMV, and then co-transfected with supercoiled pAdEasy-1, which was an adenoviral backbone vector except for deletions of E1 and E3, to competent E. coli BJ5183 for homogenous recombination using electroporation procedure. With the same method, pAdTrack-CMV was also co-transformed with pAdEasy-1 for homogenous recombination in BJ5183. Identified with restriction endonuclease Pad and polymerase chain reaction （PCR）, plasmids pAd-GFP/tk and pAd-GFP were successfully constructed. Each of them was digested with Pacl and sequently transfected into human embryo kidney 293 cells （HEK293） using Lipofectamine 2000. Results Comet-like adenovirus-producing foci of Ad-GFP/tk and Ad-GFP were observed after 5 to 7 days of cell culture After twelve days of packaging, the replication-defective adenoviruses were collected. Identified with PCR, thymidine kinase gene was successfully constructed into Ad-GFP/tk. Conclusion The replication-defective adenoviruses containing thymidine kinase can be constructed more easily by homogenous recombination in bacteria than conventional techniques.
基金Supported in part by National Cancer Institute(Bethesda,MD)grants R01 NS070289(I.U.,Charles Cobbs-PI),5R03DE021758(A.B.)generous support from Russian Fund of Fundamental Research(#No 11411.0008700.13.082 and No 13411.1008799.13.120(A.Y.B.).
文摘Glioblastoma Multiforme(GBM)is a rapidly progressing brain tumor.Despite the relatively low percentage of cancer patients with glioma diagnoses,recent statistics indicate that the number of glioma patients may have increased over the past decade.Current therapeutic options for glioma patients include tumor resection,chemotherapy,and concomitant radiation therapy with an average survival of approximately 16 months.The rapid progression of gliomas has spurred the development of novel treatment options,such as cancer gene therapy and oncolytic virotherapy.Preclinical testing of oncolytic adenoviruses using glioma models revealed both positive and negative sides of the virotherapy approach.Here we present a detailed overview of the glioma virotherapy field and discuss auxiliary therapeutic strategies with the potential for augmenting clinical efficacy of GBM virotherapy treatment.
基金supported by Techpool Bio-Pharma Co.,Ltd.(grant no.AKR-S005).
文摘Objective:To investigate the anti-tumor effects of an E1B55KD-deleted oncolytic adenovirus,H101,in combination with a humanized anti-PD-1(Programmed cell death protein 1)monoclonal antibody,Camrelizumab.Methods:Anti-tumor efficacy of intratumoral injection of H101 or/and intraperitoneal injection of Camrelizumab were evaluated in an immune system humanized NOD Prkdc^(scid) Il2rg^(-/-)mice subcutaneous(S.C.)tumor model,established with human glioblastoma of unknown origin cell line U87-MG,and human bladder cancer cell line T24 and YTS-1.The mechanism by which H101 induced anti-tumor immunity were also investigated.Results:Combining H101 with Camrelizumab demonstrated more potent anti-tumor effects than monotherapy in mouse S.C.tumor model.Increased tumor-infiltrating T cells were observed in the combined treatment group.H101 infection decreased the expression of CD47 in cancer cells,thereby promoting macrophages to phagocytose cancer cells.Following the H101-mediated activation of macrophages,increased levels of cytokines,including TNF,IL-12 and IFN-γwere observed.Moreover,when induced THP-1 cells were co-cultured with H101-treated cancer cells,expression of IFN-γwas increased in T cells.Elimination of IL-12 using an anti-IL-12 antibody abolished IFN-γproduction from T cells.In addition,infection with H101 increased PD-L1 expression in YTS-1 cells.These results suggested that H101 may act synergistically to enhance the therapeutic efficacy of PD-1 blockade in cancer via suppressing CD47 signaling,which may promote macrophages to phagocytose tumor cells and activate CD8^(+)T cells.Conclusion:The combination of H101 with PD-1 blockade exhibits potential as a novel strategy for the treatment of cancer.
基金Supported by the Nonprofit Research Institutes Foundation of Fujian Province,China,No.2020R1011003 and No.2022R1012001the Talents Training Project for the Key Young Scholars of Fujian Provincial Health Commission,China,No.2021GGA056.
文摘BACKGROUND Type 1 diabetes(T1D)results from the autoimmune-mediated loss of pancreatic β-cells.Current insulin therapies offer symptomatic relief but fall short of providing a definitive cure.Islet cell transplantation,while promising,faces limitations related to donor scarcity,procedural complexities,and the necessity for long-term immunosuppression.Consequently,there is an urgent need for innovative strategies aimed at β-cell regeneration.Patient-derived induced pluripotent stem cells(iPSCs),obtained from peripheral blood mononuclear cells(PBMCs)of T1D patients,hold great potential as a source of cells for therapeutic purposes.Therefore,the differentiation of T1D-iPSCs into functional pancreatic β-cells is a critical step toward effective β-cell replacement therapy.AIM To assess the potential of patient-derived T1D-β-like cells(differentiated from T1D-iPSCs reprogrammed from T1D-PBMCs)for restoring β-cell function in T1D.METHODS T1D-iPSCs were reprogrammed from T1D-PBMCs using an episomal vectorbased approach.Pluripotency was confirmed by flow cytometry(FCM),quantitative real-time polymerase chain reaction,genomic stability analysis,and teratoma formation assays.Differentiation into pancreatic β-cells was optimized using triiodothyronine(T3),vitamin C(Vc),and an adenovirus(M3C)encoding pancreatic duodenal homeobox-1,neurogenin 3(Ngn3),and MAF bZIP transcription factor A(MafA).Following characterization of β-cell features by immunofluorescence,quantitative real-time polymerase chain reaction,and flow cytometry,therapeutic efficacy was assessed through blood glucose monitoring after transplantation under the renal capsule of streptozotocin-induced diabetic mice.RESULTS T1D-iPSCs were successfully generated from T1D-PBMCs.These cells exhibited the hallmark characteristics of pluripotent stem cells,including appropriate morphology,differentiation potential,genomic integrity,and expression of pluripotency-associated genes.Differentiation yielded insulin-positive(insulin+)pancreatic β-like cells that,at the mRNA level,expressed key β-cell markers such as pancreatic duodenal homeobox-1,Ngn3,MafA,NeuroD,glucagon-like peptide-1 receptor,Nkx6.1,glucose transporter 2,and Kir6.2.Notably,the T3+Vc group displayed the lowest Ngn3 expression(1.31±0.38 vs 1.96±0.25 vs 2.51±0.24,P<0.01),while the M3C+T3+Vc group exhibited the highest MafA expression(0.49±0.11 vs 0.32±0.06 vs 0.29±0.08,P<0.05).Both in vitro and in vivo assessments confirmed the insulin secretion ability of the generated β-like cells;however,they did not demonstrate appropriate modulation of insulin release in response to variations in extracellular glucose concentrations.CONCLUSION T1D-iPSCs derived from T1D-PBMCs can be differentiated into insulin+β-like cells,albeit with functional immaturity.These cells represent a potential source of seed cells for β-cell replacement therapy in T1D.
基金funded by the Seed Industry Vitalization Program of Guangdong Province,grant number 2022-XPY-06–001.
文摘Fowl adenovirus(FAdV)serotype 4,recognized as the causative agent of hydropericardium syndrome(HPS)in chickens,causes substantial economic losses in poultry farming.To develop a simple,rapid,and reliable diagnostic method for the timely detection of FAdV-4 nucleic acid,we integrated the CRISPR/Cas12a system with recombinase-aided amplification(RAA).This approach enables visual detection of FAdV-4 with a sensitivity of one genome copy.The results can be obtained within 40 to 50 min without the need for complex instrumentation,making it ideal for remote field applications.Using this method,we investigated the prevalence of FAdV-4 in both common farm poultry and wild birds.Our results indicated that the FAdV-4-positive rate in wild birds was 51.19%,suggesting that wild birds may serve as specific reservoirs for this virus.In summary,we present a sensitive,swift,accurate,and inexpensive detection method for FAdV-4,along with an investigation of its epidemic situation in birds.Our study advances the detection and epidemiological understanding of FAdV-4 transmission among farm poultry and wild birds.
基金supported by the projects of National Virus Resource Center(NVRC-PY-03,E1YZ020501)Natural Science Foundation of Hubei Province(2022CFB564)+4 种基金Foundation of Hubei Provincial Health Commission(WJ2023M108)National Basic Science Data Sharing Platform(no.2018ZX10101004)National Basic Science Data Sharing Service Platform(no.NBSDC-DB-13)the International Cooperation Base of Hubei Province for Infection and Immunitysupported by Outstanding Medical Young Scholars of Hubei Province and Wuhan Young and Middle-aged Medical Backbone Talent Program.
文摘Acute respiratory tract infections(ARTIs)are among the leading causes of morbidity and mortality in children worldwide.Human adenovirus(HAdV)infections are estimated to account for at least 5%of pediatric ARTIs.The circulated genotypes of HAdV and the correlation between genotype and clinical manifestations in Wuhan,China,before and after the complete relaxation of nonpharmaceutical interventions against severe acute respiratory syndrome coronavirus 2,remain unknown.Here,101 HAdV strains were isolated from throat swab samples collected from hospitalized children with ARTIs who tested positive for HAdV nucleic acid.Of these,sixty-six strains from 2022 to twenty-three strains from 2023 were successfully genotyped and subjected to phylogenetic analysis based on the hexon,penton base,and fiber genes.Six genotypes,B3,C1,C2,C5,C104,and C108 were identified.HAdV-B3(84.85%)was the most prevalent type in 2022,while HAdV-C(86.96%),including C1,C2,C108,and C104,was the most prevalent in 2023.These strains were phylogenetically related to strains from Japan,China,and the United States in recent years.When comparing clinical characteristics,pediatric patients infected with B3,C1,C2,C5,C104,or C108 exhibited similar clinical manifestations,primarily fever and cough,but varying interleukin(IL)-10 levels.In conclusion,from June 2022 to September 2023,the circulated genotypes of HAdV in Wuhan included B3,C1,C2,C108,C5,and C104.The endemic pattern of HAdV in Wuhan,China,shifted from species B as the dominant type in 2022 to species C in 2023.
基金Scientific Research Project of Hebei Province Administration of Traditional Chinese Medicine:to Explore the Protective Effect and Mechanism of Zexie Decoction on Lipotoxic Cardiomyopathy based on the p-mitogen-activated protein kinases/Peroxisome proliferator-activated receptorγcoactivator 1-alpha(p MAPK/PGC-1α)Signaling Pathway(No.2022096)Medical Science Research Project of Hebei Province:the Effect of 23-acetyl Alismol-B on Mitochondrial Function in Palmitic Acid-induced H9c2 Cells Was Investigated based on the Ca2+-Cyclic Adenosine Monophosphate(c AMP)-Response Element Binding Protein/c AMP Response Element(CREB/CRE)-PGC-1αSignaling Pathway(No.20221490)+1 种基金Hebei province natural science fund project:Study on the Mechanism of Danshen Zexie Decoction in Activating Nuclear Factor Erythroid 2-related Factor 2 Signaling Pathway to Trigger 0mi/Htr A2,Restoring Autophagic Flux and Enhancing Metabolism-Related Fatty Liver Disease(No.H2023423064)Hebei graduate student innovation ability funding training project:to Investigate the Protective Effects and Underlying Mechanisms of Zexie Decoction on Lipotoxic Cardiomyopathy,with A Focus on the PGC-1a Signaling Pathway(No.CXZZBS2022096)。
文摘OBJECTIVE:To investigate the mechanism of Dan Ze mixture(丹泽合剂,DZM)in the treatment of lipotoxic cardiomyopathy.METHODS:Ultra-performance liquid chromatography tandem mass spectrometry was employed to characterize the serum migration constituents of DZM.A lipotoxic cardiomyopathy rat model was established through high-fat diet and intervened by different doses of DZM.The cardiac function was assessed using echocardiography,and hematoxylin and eosin,oil red O,and Masson staining were conducted to evaluate morphological changes,lipid accumulation,and fibrosis in myocardial tissue.Serum myocardial enzyme activity,lipid levels,and lipid content of myocardial tissue were measured,while fluorescent staining and colorimetry were used to assess oxidation levels in myocardial tissue.Mitochondrial membrane potential was detected by 5,5',6,6'-Tetrachloro-1,1',3,3'-tetraethyl-imidacarbocyanineiodide(JC-1).Transmission electron microscopy was employed to observe ultrastructure and mitochondrial structure changes in myocardial tissue.Fluorescence double staining and colocalization were utilized to observe the binding of autophagosomes and mitochondria,while immunohistochemical staining was used to detect the expression of mitophagy-related proteins.Terminal deoxynucleoitidyl transferase mediated nick end labeling staining was employed for the identification of apoptosis in myocardial tissue,while quantitative real-time reverse transcriptase polymerase chain reaction(q RT-PCR)and Western blot were utilized for the detection of apoptosis,B-cell lymphoma-2 adenovirus E1B 19 k Da-interacting protein 3(BNIP3)/mitophagy signaling pathway-related genes and proteins.In palmitic acid-induced Rat H9C2 cardiomyocytes(H9c2)cells,various cellular parameters including cell viability,lactate dehydrogenase release,apoptosis rate,oxidative stress level,mitochondrial structure and function,and mitophagy level were assessed after the treatment of DZM drug-containing serum for a duration of 24 h.The cellular expressions of BNIP3/mitophagy signaling pathway relevant genes and proteins were further evaluated using q RT-PCR and Western blot techniques.RESULTS:A total of 295 prototypes(e.g.,phenolic acids,quinones,terpenoids)were identified in serum of rats after oral administration of DZM.In vivo,DZM therapy has been shown to effectively enhance cardiac function,mitigate high-fat diet-induced myocardial structural damage and lipid accumulation.Furthermore,DZM has demonstrated the ability to reduce lipid levels,attenuate cell apoptosis,combat oxidative stress,enhance mitochondrial structure and function,and activate the BNIP3/mitophagy signaling pathway.Furthermore,the silencing of BNIP3 has been shown to exacerbate palmitic acid-induced damages in H9c2 cells,while inhibiting the BNIP3/mitophagy signaling pathway can mitigate the inhibitory effects of DZM on palmitic acidinduced apoptosis,lipid deposition and oxidative stress.CONCLUSION:This study presents preliminary evidence for the therapeutic efficacy of DZM on lipotoxic cardiomyopathy through the activating BNIP3/mitophagy signaling pathway.
基金supported by National Key Research and Development Program of China(2023YFC2306001)National Natural Science Foundation of China(No.32470141)+1 种基金the Beijing Research Center for Respiratory Infectious Diseases Project(BJRID2025-008)CAMS Innovation Fund for Medical Sciences(CIFMS,NO.2019-I2M-5–026,2022-I2M-CoV19-006).
文摘Human adenovirus type 108(HAdV-108)has been detected in multiple countries,including China,and is associated with severe acute respiratory infection(ARI)in children,with reported fatalities.However,studies on HAdV-108 remain limited.This study aimed to investigate the clinical and genetic characteristics of HAdV-108 in ARI children in China.From 2014 to 2024,6720 respiratory samples were collected from hospitalized children with ARI at ten hospitals across northern and southern China,of which 505(7.51%)tested positive for HAdV.The whole-genome and three major capsid protein genes were amplified and sequenced for bioinformatics analysis,which revealed that among 317 HAdV-isolated samples,21(6.62%)were identified as HAdV-108,ranking third after HAdV-114 and HAdV-7.Clinical analysis of HAdV-108-positive cases showed that the main manifestations were cough and fever.Seven children had gastrointestinal symptoms,and two children without underlying diseases were diagnosed with severe pneumonia.Phylogenetic analysis of wholegenome sequences revealed distinct predominant epidemic branches between domestic and international strains,with one strain obtained in this study forming an independent branch.Hexon protein exhibited the fastest evolution rate,lowest identity,and greatest amino acid variability,while fiber protein displayed the slowest evolution rate,highest identity,and greatest conservation and stability.Compared with the earliest reported HAdV-108 strain,three amino acid deletions were identified in the RGD loop region of penton base protein,resulting in potential structural change.Recombination analysis identified five distinct recombination patterns.In vitro experiments demonstrated that HAdV-108 had proliferation capacity comparable to other species C adenoviruses.In summary,HAdV-108 has persistently circulated in China,causing severe ARIs and concurrent gastrointestinal manifestations.Cluster3 was the predominant epidemic branch in China.HAdV-108 exhibited significant intratype genetic variation,with random and diverse recombination events.
文摘BACKGROUND Congenital short bowel syndrome(CSBS)is a rare disorder characterized by a congenital shortage in the length of the small intestines,resulting in compromised intestinal functionality,frequently accompanied by congenital intestinal malrotation.This study summarizes the experience of enteral and parenteral nutrition(PN)schemes for two cases of CSBS accompanied by intestinal malrotation to provide a reference for this condition.CASE SUMMARY Case 1 underwent surgical intervention 23 days after birth,but experienced postoperative intolerance to oral feeding.Consequently,the patient was transitioned to nasal feeding for 6 months while concurrently receiving round-the-clock PN.Despite ongoing adjustments to the caloric intake of enteral and PN throughout the treatment,cholestatic liver injury persisted.Furthermore,this child developed malnutrition by the age of 1 year.Case 2 received surgical intervention on 2 days after birth,demonstrating favorable tolerance for enteral feeding and intermittent PN.The child exhibited satisfactory growth and development without any associated complications.CONCLUSION CSBS is rarely encountered in clinical practice and is often accompanied by congenital intestinal malrotation.It can be improved with early diagnosis and active surgical intervention.Thus,we recommend early initiation of total PN,which is a highly effective measure in promoting early growth and development.
基金Support by Science and Technology Development Program of Jilin Province(20080106)~~
文摘[Objective] Sheep contagious ecthyma virus B2L gene recombinant adenovirus was built by adenovirus vector system.[Method] Genome DNA extracted from sheep contagious ecthyma virus strain JLSY04 as a template,Gene fragments obtained from B2L by PCR amplification;B2L gene cloning was cloned into PDNR-CMD vector,screening positive clones and plasmid CTC572-6 was obtained;CTC572-6 plasmid for homologous was recombined with the adenoviral vector.Screening positive clones and bacilli PCR,digestion and sequencing and so on were identified.[Result] After identified by enzyme digestion and gene sequencing,recombinant adenovirus vector CTC572Ade-30 of carrying sheep contagious ecthyma virus B2L gene was constructed successfully.[Conclusion] Which laid the foundation for sheep contagious ecthyma genetically engineered vaccine.
基金This work is supported by International Cooperation Important Project of National Natural Science Foundation of China(No.30120160824)the State 863 High Technology R&D Project of China(No.2001AA217031).
文摘Objective: To evaluate the therapeutic efficacy of replicative adenovirus CNHK500 in the treatment of hepatocellular carcinoma. Methods: Virus proliferation assay, cell viability assay and Western blot were performed to assess the selective replication and cytolysis of CNHK500 in telomerase positive liver cancer cells Hep3B, HepGII, SMMC7721 and in normal cells. Results: The replicative multiples of CNHK500 in HepGII, Hep3B and SMMC7221 after 96 h of virus proliferation were 52 000, 396 984.9 and 632 911.3 fold respectively, similar to those of wtAd5. However, CNHK500 demonstrated more significant attenuated replicative ability in normal cell lines than wtAd5. CNHK500 replicated only 3.1-100 fold at 96 h, while the wtAd5 still reached 3160-17 357 fold. CNHK500 could cause half of HepGII cells death within 7 days at MOI 2, in Hep3B cell lines the IC50 was as low as MOI 0.01, whereas the IC50 in BJ cell was as high as MOI 1000. CNHK500 E1A protein could only be detected in hepatocellular cancer cells but not in normal cells under normoxia. E1B protein could only be detected under hypoxia condition at a MOI of 1. Conclusion: CNHK500 can efficiently replicate in and kill liver cancer cells as well as wtAd5 do while it is severely attenuated in proliferation and cytolysis among normal cells. It would be a prominsing strategy for liver cancer tratment.
基金supported by Key Project of Qinghai Health and Family Planning Commission [2017-wjzd-08]Qinghai Thousand People Plan,Qinghai High-level Talents Plan in Public Health and National Science and Technology Major Project of the Ministry of Science and Technology of China during ‘13th Five-Year Plan’[2017zx10004-208]
文摘Human adenoviruses (HAdVs) belong to the genus Mastadenovirus, family Adenoviridae, and can cause respiratory tract and gastrointestinal tract infections, as well as ocular infections in humans[1]. Till date, a total of 84 unique genotypes of AdVs have been identified and classified as human Mastadenovirus species A to G, and specific types are often associated with certain clinical symptoms, epidemiological settings, and demographic risk groups[2]. Among these species, members of the species HAdV-B (HAdV-3, HAdV-7, HAdV-11, HAdV-16, HAdV-21, HAdV-34, HAdV-35, HAdV-50, etc), HAdV-C (HAdV-1, HAdV-2, HAdV-5, and HAdV-6), and HAdV-E (HAdV-4) have been generally associated with respiratory infections[3]. HAdV infections are mild and self-limited in healthy individuals, whereas they can result in high mortality rates in immunocompetent and immunocompromized patients[4].
基金Supported by Maurits en Anna de Kock fund to MA van Geer
文摘AIM:To generate an adenoviral vector specifically targeting the EphA2 receptor(EphA2R) highly expressed on pancreatic cancer cells in vivo.METHODS:YSA,a small peptide ligand that binds the EphA2R with high affinity,was inserted into the HI loop of the adenovirus serotype 5 fiber knob.To further increase the specificity of this vector,binding sites for native adenoviral receptors,the coxsackie and adenovirus receptor(CAR) and integrin,were ablated from the viral capsid.The ablated retargeted adenoviral vector was produced on 293T cells.Specifi c targeting of this novel adenoviral vector to pancreatic cancer was investigated on established human pancreatic cancer cell lines.Upon demonstrating specifi c in vitro targeting,in vivo targeting to subcutaneous growing human pancreatic cancer was tested by intravenous and intraperitoneal administration of the ablated adenoviral vector.RESULTS:Ablation of native cellular binding sites reduced adenoviral transduction at least 100-fold.Insertion of the YSA peptide in the HI loop restored adenoviral transduction of EphA2R-expressing cells but not of cells lacking this receptor.YSA-mediated transduction was inhibited by addition of synthetic YSA peptide.The transduction specificity of the ablated retargeted vector towards human pancreatic cancer cells was enhanced almost 10-fold in vitro.In a subsequent in vivo study in a nude(nu/nu) mouse model however,no increased adenoviral targeting to subcutaneously growing human pancreas cancer nodules was seen upon injection into the tail vein,nor upon injection into the peritoneum.CONCLUSION:Targeting the EphA2 receptor increases specificity of adenoviral transduction of human pancreatic cancer cells in vitro but fails to enhance pancreatic cancer transduction in vivo.
